Electrophysiological and molecular characterization of neurons in the nucleus of the solitary tract after cholinergic activation

Abstract

The nucleus of the solitary tract (NTS) is the first projection site of afferent fibers from peripheral arterial baroreceptors and chemoreceptors. Previous studies have identified a cholinergic system in the NTS. Recent studies from our laboratory have shown that increasing doses of acetylcholine (ACh, 1, 10 and 100 mM) injected into the intermediate portion of the NTS (iNTS) in rats submitted to the in situ working heart-brainstem-hypothalamus preparation induces inhibition responses of sympathetic nerve activity (SNA) and reduction of the phrenic nerve depolarization frequency (PNA). When injected into the commissural portion of the NTS (cNTS), ACh did not cause significant changes in SNA, but promoted an increase in PNA. Furthermore, ACh injected into the cNTS promoted a change in the sympathetic-respiratory coupling pattern. It was also observed that the ACh-induced tachypnea in cNTS was reduced by pre-treatment with atropine (non-selective muscarinic antagonist, 17 mM). On the other hand, atropine did not promote significant changes in the tachypnea and sympatoexcitation magnitudes induced by chemoreflex with KCN. The administration of mecamylamine (non-selective nicotinic antagonist, 5 mM) into the cNTS decreased the ACh-induced tachypnea and KCN-induced tachypnea. On the other hand, mecamylamine did not significantly change the KCN-induced sympatoexcitation magnitude. These results suggest that ACh plays an important role in controlling cardiovascular and respiratory efferent activities, with distinct roles between iNTS and cNTS, involving, at least in cNTS both muscarinic and nicotinic receptors. Previous studies, in experiments performed on brainstem horizontal slices, it was demonstrated that iNTS neurons have an increase in the frequency of the action potentials induced by ACh. However there are no studies so far on the electrophysiological effects of ACh into the different NTS subnuclei, specifically in the cNTS. Considering that iNTS and cNTS show distinct responses on SNA and PNA to ACh, it would be interesting to verify whether ACh has differential effects in terms of changes in action potentials frequency and membrane voltage of neurons in these subnuclei. Recent studies from Dr. Ferguson's laboratory have made an association between electrophysiological studies (patch clamp) and molecular characterization (mRNA) of the recorded neurons, using the single cell RT-PCR technique. Therefore, the present study we aim to investigate the electrophysiological and molecular properties of iNTS and cNTS neurons by cholinergic activation with ACh. (AU)